JP6884608B2 - Compositions and external preparations for skin - Google Patents

Description

The present invention is a functional material having a plurality of natural product-derived components and having excellent physiological activity and biosafety, a skin (including scalp) external preparation containing the same, and food and drink for beauty or health promotion. It is a product.

In recent years, research has been conducted on cell aging and damage to cells caused by external factors (for example, ultraviolet rays, chemical substances such as air pollutants and environmental hormones, allergens such as pollen, environmental stress, etc.), and various cells have been studied. The mechanism of senescence and cell damage and repair has been elucidated.

For example, regarding skin cells, internal factors such as inactivation of cell proliferation / differentiation with aging, decrease in hormone secretion, quantitative decrease in extracellular matrix components (collagen, laminin, etc.) constituting the skin, and the sun Aging phenomenon (wrinkles, tarmi, etc.) due to complex intertwining of active oxygen induced by light (ultraviolet rays), chemical substances such as air pollutants and environmental hormones, allergens such as pollen, and external factors such as environmental stress. It is known that rough skin and changes in color tone occur.

Furthermore, ultraviolet rays, chemical substances, and allergens, which are external factors, damage cells and tissues in the living body to alter biological components or generate active oxygen. As a result, an antigen is generated inside the cell, and inflammation occurs in the living body. Furthermore, the above-mentioned external factors induce abnormal deposition of intracellular melanin pigment, causing spots, freckles, liver spots and the like on the skin.

For the purpose of preventing and improving the unhealthyness and aging of cells as described above, various active ingredients have been conventionally proposed, and cosmetics, foods and drinks and pharmaceuticals containing these active ingredients have been put on the market. For example, antioxidants such as vitamin C, vitamin E and catalase; anti-inflammatory agents such as glycyrrhizic acid or salts thereof, allantin, tranexamic acid; various ultraviolet absorbers; α-hydroxycarboxylic acid, placenta extract, γ-amino- Cell-activating components such as β-hydroxybutyric acid; extracellular matrix components such as collagen, elastin, hyaluronic acid or salts thereof; moisturizers such as urea; protein glycation inhibitors such as aminoguanidine. Further, kojic acid, linoleic acid and the like are known as substances that suppress the occurrence of pigmentation such as skin stains, freckles and chloasma, and are widely used as active ingredients of whitening agents.

As described above, cell activators, anti-aging agents and whitening agents have been conventionally proposed based on the mechanism of cell aging phenomenon and unhealthyness, but they are safe for living organisms and actually applied to living organisms. There is a problem in terms of effectiveness when applied or taken. Therefore, there is a demand for a functional material in which these conventional problems are improved.

In view of the problems of the prior art, the present inventors have conducted diligent research to find a new active ingredient derived from a natural product from the viewpoint of biosafety. As a result, by the extract of Plum belonging to the genus Plum of the Rosaceae family and the extract of any one or more of yeast, aspergillus and lactic acid bacteria, or the microorganism of any one or more of yeast, aspergillus and lactic acid bacteria. The composition containing the produced microbial-derived fermentation metabolite has excellent cell activation effect, antioxidant effect, collagen synthesis promoting effect, melanin synthesis suppression and fat accumulation suppression, and is used as an external preparation for skin (including scalp) and food and drink. We have found that it is useful as a functional material for goods.

Conventionally, it is disclosed in Patent Documents 1 to 6 that an extract of a cherry or a yeast extract belonging to the genus Plum of the Rosaceae family has skin physiological activity. A composition containing an extract of any one or more of the above or a microbial-derived fermented metabolite produced by any one or more of yeast, aspergillus and lactic acid bacteria, can be used as an external preparation for skin or eat and drink. It was not known that it was useful as a functional material for goods.
Japanese Patent Application Laid-Open No. 63-060935 Japanese Patent Application Laid-Open No. 01-090131 Japanese Patent Application Laid-Open No. 06-279256 Japanese Unexamined Patent Publication No. 58-172321 Japanese Unexamined Patent Publication No. 11-106336 Japanese Unexamined Patent Publication No. 64-003126

The present invention relates to an extract of Plum belonging to the genus Plum of the Rosaceae family, an extract of any one or more of yeast, aspergillus and lactic acid bacteria, or a microorganism of any one or more of yeast, aspergillus and lactic acid bacteria. It is a composition containing a fermentation metabolite derived from a microorganism produced by.
The present invention is also characterized in that it is a skin (including scalp) external preparation or food or drink containing the above composition.

The present invention relates to an extract of cherry blossoms belonging to the genus Sakura of the family Rose family, an extract of any one or more of yeast, aspergillus and lactic acid bacteria, or a microorganism of any one or more of yeast, aspergillus and lactic acid bacteria. It is a composition containing a fermentation metabolite derived from a microorganism produced by the above, and the composition has a synergistic effect of cell activation, antioxidant action, promotion of collagen production, suppression of melanin synthesis and suppression of fat accumulation. , Cell health effect, prevention and improvement effect of damage to living body (oxidative damage and inflammatory damage) due to external factors such as ultraviolet rays, moisturizing effect, skin turnover improvement effect, prevention and improvement of skin wrinkles and melanin It exerts effects, prevention and improvement effects of spots and buckwheat, sebum suppression effect and slimming effect. As a result, the composition according to the present invention is useful as a functional material for external preparations for skin and foods and drinks. Furthermore, since it also has the effect of promoting the production of collagen in the scalp, it is useful as a functional material for preventing hair loss or gray hair.

Hereinafter, preferred embodiments of the present invention will be described in detail.
The present invention is derived from an extract of cherry blossoms and an extract of any one or more of yeast, aspergillus and lactic acid bacteria, or a microorganism produced by any one or more of yeast, aspergillus and lactic acid bacteria. It is a composition containing a fermentation metabolite.

In the present invention, "Sakura" is a plant belonging to the genus Plum of the Rosaceae family, for example, Yamazakura (Prunus jamasakura) such as Oshimazakura, Oyamazakura, Kasumizakura, Yamazakura (Yoshino cherry tree), Yaezakura, Kanzan, Fugenzo, Yoshino cherry tree, Examples thereof include Satozakura (Prunus lannesiana) such as Ichiyo, Someiyoshino (Prunus × yedoensis), Kanhizakura, Edohigan, Mamezakura, Choujizakura, etc., but the cherry tree of the present invention is not limited thereto. Further, as the site of use, any of bark, flowers, seeds, fruits, leaves, roots and the like may be used, but the use of bark or flowers is preferable.

To prepare the extract of cherry blossoms, first, the site to be used is washed with water in advance to remove foreign substances, if necessary, and then left as it is or dried, and if necessary, shredded or crushed and brought into contact with an extraction solvent. Extract. Extraction can be carried out by contacting with an extraction solvent according to a conventional method such as a dipping method, but a steam distillation method or a supercritical extraction method can also be used in addition to the dipping method.

As the extraction solvent, water; lower alcohols such as methanol, ethanol and propanol; polyhydric alcohols such as ethylene glycol, 1,2-propanediol, 1,3-propanediol, 1,3-butylene glycol and glycerin; Esters such as ethyl acetate, butyl acetate and methyl propionate; ketones such as acetone and methyl ethyl ketone; ethers such as ethyl ether and isopropyl ether; hydrocarbon solvents such as n-hexane, toluene and chloroform are included. They can be used alone or in admixture of two or more.

Among the extraction solvents, hydrophilic solvents such as water, lower alcohols and polyhydric alcohols are preferable. Preferred examples of using this hydrophilic solvent include, for example, water, lower alcohols (particularly ethanol), or polyhydric alcohols (particularly 1,2-propanediol, 1,3-propanediol, 1,3-. Butylene glycol) can be used alone, or a mixed solvent of water and lower alcohols, or a mixed solvent of water and polyhydric alcohols can be used. Among them, water alone or water and 1,2- A mixed solvent of propanediol, 1,3-propanediol or 1,3-butylene glycol is particularly preferable.

When a mixed solvent is used, for example, in the case of a mixed solvent of water and 1,2-propanediol, 1,3-propanediol or 1,3-butylene glycol, the volume ratio (hereinafter the same applies) is 1. : 5 to 20: 1, preferably in the range of 1: 5 to 25: 1 for a mixed solvent of water and ethanol, and preferably in the range of 1: 10 to 20: 1 for a mixed solvent of water and glycerin. ..

The weight ratio of the site where the cherry tree is used and the extraction solvent is preferably 1: 1 to 1: 100, more preferably 1: 5 to 1:60.

The pH at the time of preparation of the extract is not limited, but is generally preferably in the range of 3 to 9. In this sense, if necessary, an alkali adjusting agent such as sodium hydroxide, sodium carbonate, potassium hydroxide or the like, or an acid adjusting agent such as citric acid, hydrochloric acid, phosphoric acid, sulfuric acid, etc. is added to the extraction solvent, which is desired. The pH may be adjusted to the above.

Extraction conditions such as extraction temperature and extraction time vary depending on the type of solvent used and pH, but for example, water or 1,3-butylene glycol, or water and 1,2-propanediol, 1,3-propanediol or When a mixed solution with 1,3-butylene glycol is used as a solvent, the extraction temperature is preferably in the range of 0 ° C. to 90 ° C., and the extraction time is preferably 1 to 168 hours (1 hour to 1 week). It is more preferably in the range of 1 to 120 hours (1 hour to 5 days).

If necessary, the extract may be hydrolyzed prior to the extraction treatment of the present invention or in parallel with the extraction treatment. As a result, there is a possibility that the storage stability of the extract can be improved and the extract as a functional material for external preparations for skin and foods and drinks can be used more effectively.

When the extract is subjected to enzyme hydrolysis treatment, the enzymes include proteolytic enzymes such as actinase, papaine and pepsin, starch degrading enzymes such as glucoamylase, α-amylase and β-amylase, cellulase, hemicellulase and pectinase. One or more selected from any of the enzyme groups such as fibrinolytic enzyme and lipid degrading enzyme such as lipase may be used, but one or more selected from those enzyme groups, respectively. It is more preferable to use the above enzymes in combination.

The amount of the enzyme added is preferably in the range of 0.00001 to 10% by weight in total with respect to the solid content of the site where the plant is used.

In the present invention, yeast is, for example, Saccharomyces cerevisiae, Saccharomyces awamori, Saccharomyces chevalieri, Saccharomyces cerevisiae, Saccharomyces calsbergensis, Saccharomyces carlsbergensis, Saccharomyces carlsbergensis, Saccharomyces carlsbergensis, Saccharomyces carlsbergensis. Yeasts of the genus Galactomyces, yeasts of the genus Galactomyces, yeasts of the genus Torulaspora delbruekii, Torulaspora fermentati, Torulaspora rosei, etc. Zygosaccharomyces soya, Zygosaccharomyces sake, Zygosaccharomyces miso, Zygosaccharomyces lactis, Zygosaccharomyces lactis, Zygosaccharomyces lactis, etc. ), Candida kefyr, Candida sake, Candida scottii and other Candida yeasts, Aureobasidium Pullulans, Aureobasidium mansonii , Aureobasideium microstictum and other yeasts of the genus Aureobasideium. Further, the yeast according to the present invention may be any of sake yeast, wine yeast, brewer's yeast, yeast derived from plant flowers (rose, lily, cherry tree, etc.), and yeast derived from the sea.

In the present invention, examples of aspergillus include Aspergillus oryzae, Aspergillus flavus, Aspergillus polyoxogenes, Aspergillus sojae, Aspergillus aspergillus, Aspergillus aspergillus, and Aspergillus aspergillus. Aspergillus kawauchii, Aspergillus usami, Aspergillus niger and other black aspergillus, Monascus anka, Monascus pilosus and other red aspergillus.

In the present invention, the lactic acid bacterium is, for example, a lactic acid bacterium of the genus Lactobacillus delbrueckii such as Lactobacillus plantarum, L. brevis, Lactobacillus casei, and Lactobacillus delbrueckii. Lactic acid bacteria of the genus Carnobacterium such as Carnobacterium divergens and Carnobacterium piscicola; Leuconostoc mesenteroides, Leuconostoc citreum, etc. Lactic acid bacteria of the genus Leuconostoc; Lactic acid bacteria of the genus Streptococcus such as Streptococcus faecalis and Streptococcus pyogenes; Lactic acid bacteria of the genus; Lactococcus plantarum Lactococcus rafinolactis and other lactic acid bacteria of the genus Lactococcus; Weissella confusa, Weissella kandleri and other lactic acid bacteria of the genus Weissella kandleri (Atopobium minutum), Atopobium parvulus and other lactic acid bacteria of the genus Atopobium; Vagococcus fluvialis, Vagococcus fluvialis, Vagococcus salmoninarum, etc. Examples thereof include lactic acid bacteria belonging to the genus Pediococcus such as Su (Pediococcus damnosus) and Pediococcus pentosaceus.

In the present invention, an extract of a microorganism can be obtained by culturing yeast, aspergillus or lactic acid bacterium in a medium and collecting the culture solution by an operation such as filtration. In addition, a fermented product obtained by fermenting a plant such as rice, adlay, soybean, etc. using any one or more of these microorganisms may be used as a fermentation metabolite derived from a microorganism as an active ingredient of the composition according to the present invention. In this case, as the microbial-derived fermentation metabolite, the solid content phase in the fermented product may be used, or a fermented liquid obtained by removing the solid content from the fermented product may be used.

Further, in the present invention, as a fermentation metabolite derived from microorganisms, brewed liquor or distilled liquor produced by using yeast, aspergillus and / or lactic acid bacterium, or residue (sake lees, liquor lees) produced in the production process of brewed liquor or distilled liquor. , Wine lees, beer lees) and extracts obtained by extraction treatment from their residues can also be used.

The composition of the present invention can be incorporated into, for example, skin (including scalp) external preparations (cosmetics, quasi-drugs, external medicines), and foods and drinks for beauty or health promotion. Examples of external skin preparations include emulsions, creams, lotions, essences, packs, lipsticks, foundations, liquid foundations, make-up press powders, blushers, white powder, pigment wash, body shampoos, scalp, hair shampoos, and hair conditioners. , Shampoo or tonic for hair growth and hair growth, cleansing cosmetics such as soap, and bathing agents, but the present invention is not limited thereto. In addition, as foods and drinks for beauty or health promotion, beverages such as beauty beverages, energy drinks, sports drinks, near water, vitamin beverages, mineral beverages, alcoholic beverages; various soups (including powdered soups), dairy products , Jelly, candy, tablet confectionery, gum, etc .; Although it can be blended in tablets, liquid, granular or jelly-like health foods and beverages, the present invention is not limited to these, and foods and drinks that can be taken orally. Can be blended in products, etc.

The blending amount of the composition of the present invention shall be in the range of 0.002 to 1.0% by weight (solid content weight%, the same shall apply hereinafter) in the case of basic cosmetics as the solid content of the composition, and of makeup cosmetics. In the case, it is in the range of 0.002 to 1.0% by weight, and in the case of cosmetics for cleaning, it is in the range of 0.002 to 10.0% by weight. In the case of hair cosmetics, the solid content of the composition is in the range of 0.0001 to 5.0% by weight. The blending amount of the composition of the present invention in foods and drinks is preferably in the range of 0.1 to 15% by weight as the solid content of the composition.

When the composition of the present invention is blended with an external preparation for skin or food and drink, in addition to the composition which is an essential component, components usually used in those products, such as an oil component and a surfactant (synthetic system, natural product system) ), Moisturizer, thickener, preservative / bactericidal agent, powder component, ultraviolet absorber, antioxidant, pigment, fragrance and the like can be appropriately blended as needed. Further, as long as the effectiveness and features of the composition are not impaired, other physiologically active ingredients may be combined and blended at all.

Oily components include, for example, lotus oil, olive oil, jojoba oil, castor oil, soybean oil, rice oil, rice bran oil, rice germ oil, palm oil, chamomile oil, palm oil, cacao oil, meadowfoam oil, rosehip oil, etc. Rose oil, lanbender oil, sheer butter, tea tree oil, avocado oil, macadamia nut oil, bergamot oil, plant-derived oils and fats such as squalane; animal-derived oils and fats such as mink oil and turtle oil; , Rice wax, waxes such as lanolin; hydrocarbons such as liquid paraffin, vaseline, paraffin wax, squalane; fatty acids such as myristic acid, palmitic acid, stearic acid, oleic acid, isostearic acid, eicosenoic acid; lauryl alcohol, Higher alcohols such as cetanol and stearyl alcohol; synthetic esters such as isopropyl myristate, isopropyl palmitate, butyl oleate, 2-ethylhexyl glyceride, higher fatty acid octyldodecyl (octyldodecyl stearate, etc.) and synthetic triglycerides, etc. Be done.

Examples of the surfactant include polyoxyethylene alkyl ether, polyoxyethylene fatty acid ester, polyoxyethylene sorbitan fatty acid ester, glycerin fatty acid ester, polyglycerin fatty acid ester, polyoxyethylene glycerin fatty acid ester, polyoxyethylene hydrogenated castor oil, and poly. Nonionic surfactants such as oxyethylene sorbitol fatty acid esters; fatty acid salts, alkyl sulfates, alkylbenzene sulfonates, polyoxyethylene alkyl ether sulfates, polyoxyethylene fatty amine sulfates, polyoxyethylene alkylphenyl ether sulfates, Anionic surfactants such as polyoxyethylene alkyl ether phosphates, α-sulfonated fatty acid alkyl ester salts, polyoxyethylene alkyl phenyl ether phosphates; quaternary ammonium salts, primary to tertiary fatty amine salts, tri Cationic surfactants such as alkylbenzylammonium salt, alkylpyridinium salt, 2-alkyl-1-alkyl-1-hydroxyethylimidazolinium salt, N, N-dialkylmorphonium salt, polyethylene polyamine fatty acid amide salt; N, N-dimethyl-N-alkyl-N-carboxymethylammoniobetaine, N, N, N-trialkyl-N-alkylene ammoniocarboxybetaine, N-acylamide propyl-N', N'-dimethyl-N'- An amphoteric surfactant such as β-hydroxypropylammoniosulfobetaine can be used.

Examples of emulsifiers or emulsifying aids include stevia derivatives such as enzyme-treated stevia, saponins or derivatives thereof, casein or salts thereof (sodium, etc.), sugar-protein complexes, sucrose or esters thereof, lactose, and water-soluble soybeans. Polysaccharide, complex of soybean-derived protein and polysaccharide, lanolin or its derivative, cholesterol, stevia derivative (stevia enzyme treated product, etc.), silicate (aluminum, magnesium, etc.), carbonate (calcium, sodium, etc.), saponin and its Derivatives, lecithin and its derivatives (hydrogenated lecithin, etc.), lactic acid bacteria fermented rice, lactic acid bacteria fermented sprouted rice, lactic acid bacteria fermented grains (wheat, beans, miscellaneous grains, etc.) and the like can also be blended.

Examples of the moisturizing agent include glycerin, propylene glycol, dipropylene glycol, 1,3-butylene glycol, polyethylene glycol, sorbitol, xylitol, sodium pyrrolidone carboxylate and the like, and saccharides such as trehalose and mucopolysaccharides (for example, hyalurone). Acids and their derivatives, chondroitin and its derivatives, heparin and its derivatives, etc.), elastin and its derivatives, collagen and its derivatives, NMF-related substances, lactic acid, urea, higher fatty acids octyldodecyl, seaweed extract, silane roots (white) Examples include extracts, various amino acids and their derivatives.

Examples of the thickener include brown algae such as alginic acid, agar, carrageenan, and fucoidan, components derived from green algae or red algae; silane root (white and) extract; pectin, locust bean gum, aloe polysaccharide, alkaligenes-producing polysaccharide, and the like. Polysaccharides; gums such as xanthan gum, tragant gum, guar gum; cellulose derivatives such as carboxymethyl cellulose and hydroxyethyl cellulose; synthetic polymers such as polyvinyl alcohol, polyvinylpyrrolidone, carboxyvinyl polymer, alginic acid / methacrylic acid copolymer; hyaluronic acid And derivatives thereof; examples thereof include polyglutamic acid and derivatives thereof.

Examples of antiseptic / bactericidal agents include urea; paraoxybenzoic acid esters such as methyl paraoxybenzoate, ethyl paraoxybenzoate, propyl paraoxybenzoate, and butyl paraoxybenzoate; phenoxyethanol, dichlorophene, hexachlorophene, chlorhexidine hydrochloride, and benza chloride. Plants such as luconium, salicylic acid, ethanol, undesyleneic acid, phenols, jamar (imidazodeine luurea), polyphosphate, propanediol, 1,2-pentanediol, various essential oils, dried bark, radish fermented liquid, sugar cane, etc. Derived ethanol or 1,3-butylene glycol and the like.

Examples of powder components include cericite, titanium oxide, talc, kaolin, bentonite, zinc oxide, magnesium carbonate, magnesium oxide, zirconium oxide, barium sulfate, silicic anhydride, mica, nylon powder, polyethylene powder, silk powder, and cellulose. There are system powders, grain powders (rice, wheat, corn, talc, etc.), beans (soybeans, talc, etc.) powders, etc.

Examples of the ultraviolet absorber include ethyl paraaminobenzoate, ethylhexyl paradimethylaminobenzoate, amyl salicylate and its derivatives, 2-ethylhexyl paramethoxycinnamate, octyl silicate, oxybenzone, 2,4-dihydroxybenzophenone, 2-hydroxy-4. -Methoxybenzophenone-5-sulfonate, 4-tershally butyl-4-methoxybenzoylmethane, 2- (2-hydroxy-5-methylphenyl) benzotriazole, urocanic acid, ethyl urocanate, aloe extract, etc. ..

Antioxidants include, for example, butylhydroxyanisole, butylhydroxytoluene, propyl gallate, beautyberry extract, silane root extract, shakuyaku extract, vitamin E and its derivatives (for example, vitamin E nicotinate, vitamin E linoleate, etc.) ) Etc.

Whitening agents include t-cycloamino acid derivatives, kodiic acid and its derivatives, ascorbic acid and its derivatives, hydroquinone or its derivatives, ellagic acid and its derivatives, nicotinic acid and its derivatives, resorcinol derivatives, tranexamic acid and its derivatives, 4 -Potasium salicylic acid salt, magnolignan (5,5'-dipropyl-biphenyl-2,2'-diol), 4- (4-hydroxyphenyl) -4-butanol)), hydroxybenzoic acid and its derivatives, vitamin E And derivatives thereof, α-hydroxy acid, and AMP (adenosine monophosphate, adenosine monophosphate), and these may be blended alone or in combination of two or more.

Examples of the ascorbic acid derivative include ascorbic acid esters such as ascorbic acid monobutyrate, ascorbic acid monocaplate, ascorbic acid monopalmitate, and ascorbic acid dibutyrate, ascorbic acid ethers, and ascorbic acid sugar derivatives such as ascorbic acid glucoside. However, examples of the ascorbic acid derivative include L-ascorbic acid-2-phosphate ester sodium, L-ascorbic acid-2-phosphate ester magnesium, L-ascorbic acid-2-sulfate sodium ester, and L-ascorbic acid-2. -Ascorbic acid ester salts such as magnesium sulfate, ascorbic acid sugar derivatives such as L-ascorbic acid-2-glucoside, L-ascorbic acid-5-glucoside, and 6-position acylated products of these ascorbic acid sugar derivatives (the acyl group is Hexanoyl group, octanoyl group, decanoyl group, etc.), L-ascorbic acid tetrafatty acid esters such as L-ascorbic acid tetraisopalmitic acid ester, L-ascorbic acid tetralauric acid ester, 3-O-ethylascorbic acid, L- Glucerin ascorbic acid derivatives such as ascorbic acid-2-phosphate-6-O-palmitate sodium, glyceryl ascorbic acid or an acylated derivative thereof, and bisglyceryl ascorbic acid are used as hydroquinone derivatives. Glucopyranoside), α-arbutin (hydroquinone-α-D-glucopyranoside), etc., as the tranexamic acid derivative, tranexamic acid ester (for example, tranexamic acid lauryl ester, tranexamic acid hexadecyl ester, tranexamic acid cetyl ester or a salt thereof), Examples thereof include an amide form of tranexamic acid (for example, methylamide tranexamic acid), and examples of the resorcinol derivative include 4-n-butylresorbicinol and 4-isoamylresorbic acid, and examples of the 2,5-dihydroxybenzoic acid derivative include. 2,5-diacetoxybenzoic acid, 2-acetoxy-5-hydroxybenzoic acid, 2-hydroxy-5-propionyloxybenzoic acid and the like, and nicotinic acid derivatives such as nicotinic acid amide and benzyl nicotinate are α- Examples of the hydroxy acid include lactic acid, ascorbic acid, succinic acid, citric acid, α-hydroxyoctanoic acid and the like.

Physiologically active ingredients include, for example, placenta extract, sohakuhi extract, yukinoshita extract, perilla extract, rice bran extract or its hydrolyzate, white potato extract or its hydrolyzate, white potato fermented product, shakuyaku. Extract or its hydrolyzate, Murasakikib extract, Hass seed extract or its hydrolyzate, Hass seed fermented product, Ginseng extract or its hydrolyzate, Hatomugi hydrolyzate, Royal jelly fermented product, Pandanus amalifoliaus Examples include (Pandanus amaryllifolius Roxb.) Extract, Arcangelicia flava Merrilli extract, chamomile extract and the like. In addition, coral grass extract, rice leaf extract or its hydrolyzate, eggplant (bergamot, long eggplant, Kamo eggplant, rice eggplant, etc.) extract or its hydrolyzate, apricot fruit extract, catamen giraffe, etc. Seaweed extract, extract of marine flowering plants such as Amamo, fermented soymilk, jellyfish water, rice extract or its hydrolyzate, sprouted rice extract or its hydrolyzate, black bean extract or its hydrolysis Extracts of damask rose flowers, extracts of bamboo shoots, linoleic acid and its derivatives or processed products (eg, liposome-derived linoleic acid, etc.), animal or fish-derived collagen and its derivatives, elastin and its derivatives, glycyrrhizin Acids and their derivatives (dipotassium salt, etc.), t-cycloaminomino acid derivatives, vitamin A and its derivatives, allantin, diisopropylamine dichloroacetate, γ-amino-β-hydroxybutyric acid, gentian extract, licorice extract, carrot extract, Otaneninjin extract or its fermented product, Benin extract, Mitsuishikonbu extract, Hechima extract, Anaaosa extract, Peach extract, Peach seed extract, Kiwi extract, Sunflower extract, Zizyphus joazeiro extract, Paudalco Bark extract, daily grass extract or fermented product, hibiscus flower extract or fermented product, hagoromogusa extract, cherimoya extract, mango extract, mangostin extract, funori extract, crow dragon tea extract, Benito Takashi extract , Purple orchid extract, Sansho peel or seed coat extract or hydrolyzate, Benibana flower extract, Casablanca extract, Sweet potato extract or its fermented product, Guava leaf extract, Dokudami extract, Late white yuzu extract, There are aloe extract, fig flower extract, apple extract, white asparagus extract and the like.

Next, the present invention will be described in more detail with reference to Production Examples, Formulation Examples, and Test Examples, but the present invention is not limited thereto. In the following, all parts mean parts by weight, and% means all parts by weight.

Production example 1. Preparation of composition (1) (i) Preparation of cherry extract The flowers of Satozakura (Prunus lannesiana) are dried and crushed, 900 g of purified water is added to 18 g of the crushed product, and the mixture is extracted at 80 ° C. for 2 hours and then filtered. Then, 1385 g of a brown transparent cherry extract solution was obtained (solid content concentration 0.49%).
(Ii) Microorganism-derived fermented metabolite Yeast derived from plant (Sakura) flowers is cultured in a solution containing glucose and peptone at 15 ° C. for 48 hours, and the culture solution obtained by filtering the culture solution is a yeast extract solution. (Solid content concentration 0.51%).
(Iii) Preparation of composition After mixing the cherry extract solution and the yeast extract solution at a mixing ratio of 1: 1, the insoluble matter was filtered to obtain a brown transparent extract mixed solution (solid content concentration 0. 40%)

Production example 2. Preparation of composition (2) (i) Preparation of cherry extract The flowers of Satozakura are dried and crushed, 450 g of purified water and 450 g of 1,3-butylene glycol are added to 18 g of the crushed product, and the mixture is extracted at 80 ° C. for 2 hours. Then, it was filtered to obtain 1365 g of a brown transparent cherry extract solution (solid content concentration 0.45%).
(Ii) Preparation of fermented metabolites derived from microorganisms 160 g of sake lees produced in the process of producing sake was added to 800 g of purified water, extracted at 80 ° C. for 1 hour, and then filtered to obtain 2490 g of a pale yellow transparent sake lees extract solution. (Solid concentration 0.45%)
(Iii) Preparation of composition After mixing the cherry extract and sake lees extract at a mixing ratio of 1: 9, the insoluble matter was filtered to obtain a brown transparent extract mixed solution (solid content concentration 0.34). %).

Production example 3. Preparation of composition (3) (i) Preparation of cherry extract The flowers of Satozakura are dried and crushed, 270 g of purified water and 630 g of 1,3-butylene glycol are added to 18 g of the crushed product, and the mixture is extracted at 80 ° C. for 3 hours. Then, it was filtered to obtain 1350 g of a brown transparent cherry extract solution (solid content concentration: 0.39%).
(Ii) Preparation of fermentation metabolites derived from microorganisms Lactobacillus was liquid-cultured in MRS medium at 37 ° C. for 72 hours, and the culture solution obtained by filtering the culture solution was mixed with a lactic acid bacteria extract solution (solid content concentration 0.72%). did.
(Iii) Preparation of composition After mixing the cherry extract solution and the lactic acid bacterium extract solution at a mixing ratio of 1: 2, the insoluble matter was filtered to obtain a brown transparent extract mixed solution (solid content concentration 0. 51%)

Production example 4. Preparation of Composition (4) (i) Preparation of Sakura Extract 1350 g of a brown transparent Sakura extract solution was obtained by the same method as in Production Example 1 (i) except that wild cherry was used instead of Satozakura. (Solid content concentration 0.43%).
(Ii) Preparation of microbial-derived fermented metabolites A yeast extract solution was obtained in the same manner as in (ii) of Production Example 1.
(Iii) Preparation of composition After mixing the cherry extract solution and the yeast extract solution at a mixing ratio of 1: 1, the insoluble matter was filtered to obtain a brown transparent extract mixed solution (solid content concentration 0. 37%)

Production example 5. Preparation of Composition (5) (i) Preparation of Sakura Extract 1355 g of a brown transparent Sakura extract solution was obtained by the same method as in Production Example 2 (i) except that wild cherry was used instead of Satozakura. (Solid content concentration 0.41%).
(Ii) Preparation of microbial-derived fermented metabolite A sake lees extract solution was obtained by the same method as in (ii) of Production Example 2.
(Iii) Preparation of composition After mixing the cherry extract solution and the sake lees extract solution at a mixing ratio of 1: 9, the insoluble matter was filtered to obtain a brown transparent extract mixed solution (solid content concentration 0). .32%).

Production example 6. Preparation of Composition (6) (i) Preparation of Sakura Extract 1345 g of a brown transparent Sakura extract solution was obtained by the same method as in Production Example 3 (i) except that wild cherry was used instead of Satozakura. (Solid content concentration 0.35%).
(Ii) Preparation of microbial-derived fermented metabolite A lactic acid bacterium extract solution was obtained by the same method as in (iii) of Production Example 3.
(Iii) Preparation of composition After mixing the cherry extract solution and the lactic acid bacterium extract solution at a mixing ratio of 1: 2, the insoluble matter was filtered to obtain a brown transparent extract mixed solution (solid content concentration 0. 49%)

Production example 7. Preparation of composition (7) (i) Preparation of sakura extract A brown transparent sakura extract was prepared by the same method as in Production Example 1 (i) except that Yoshino cherry tree (Prunus × yedoensis) was used instead of Satozakura. 1360 g of the solution was obtained (solid content concentration 0.42%).
(Ii) Preparation of microbial-derived fermented metabolites A yeast extract solution was obtained in the same manner as in (ii) of Production Example 1.
(Iii) Preparation of composition After mixing the cherry extract solution and the yeast extract solution at a mixing ratio of 1: 1, the insoluble matter was filtered to obtain a brown transparent extract mixed solution (solid content concentration 0. 38%)

Production example 8. Preparation of Composition (8) (i) Preparation of Sakura Extract 1355 g of a brown transparent Sakura extract solution was obtained by the same method as in Production Example 2 (i) except that Yoshino cherry tree was used instead of Satozakura. (Solid content concentration 0.40%).
(Ii) Preparation of microbial-derived fermented metabolite A sake lees extract solution was obtained by the same method as in (ii) of Production Example 2.
(Iii) Preparation of composition After mixing the cherry extract solution and the sake lees extract solution at a mixing ratio of 1: 9, the insoluble matter was filtered to obtain a brown transparent extract mixed solution (solid content concentration 0). .31%).

Production example 9. Preparation of Composition (9) (i) Preparation of Sakura Extract 1340 g of a brown transparent Sakura extract solution was obtained by the same method as in Production Example 3 (i) except that Yoshino cherry tree was used instead of Satozakura. (Solid content concentration 0.34%).
(Ii) Preparation of microbial-derived fermented metabolite A lactic acid bacterium extract solution was obtained by the same method as in (iii) of Production Example 3.
(Iii) Preparation of composition After mixing the cherry extract solution and the lactic acid bacterium extract solution at a mixing ratio of 1: 2, the insoluble matter was filtered to obtain a brown transparent extract mixed solution (solid content concentration 0. 48%)

Production example 10. Preparation of Composition (10) (i) Preparation of Sakura Extract A cherry extract solution was obtained by the same method as in Production Example 2 (i).
(Ii) Preparation of fermented metabolites derived from microorganisms After steaming and cooling 50 g of polished rice, Aspergillus oryzae was added to prepare rice jiuqu under a temperature condition of 30 ° C. To this, 50 g of separately steamed polished rice and 200 g of sterilized purified water were added, and a pre-cultured Saccharomyces cerevisiae culture solution was added, and fermentation was carried out at 30 ° C. After the fermentation was completed, this rice fermented product solution was heat sterilized at 90 ° C. for 1 hour, then returned to room temperature and filtered to obtain 126 g of a pale yellow transparent rice fermented product solution (solid content concentration: 4.00%). ).
(Iii) Preparation of composition After mixing the cherry extract and the fermented rice solution at a mixing ratio of 1: 9, the insoluble material was filtered to obtain a brown transparent extract mixed solution (solid content concentration 0. 30%).

Production example 11. Preparation of Composition (11) (i) Preparation of Sakura Extract A cherry extract solution was obtained by the same method as in Production Example 3 (i).
(Ii) Preparation of fermented metabolites derived from microorganisms 300 g of purified water sterilized in advance was added to 100 g of rice jiuqu, and the mixture was allowed to stand at 35 ° C. Then, 100 g of separately steamed polished rice and a pre-cultured Saccharomyces cerevisiae culture solution were added thereto, and fermentation was carried out at 30 ° C. After the fermentation was completed, this rice fermented product solution was heat sterilized at 90 ° C. for 1 hour, then returned to room temperature and filtered to obtain 183 g of a pale yellow transparent rice fermented product solution (solid content concentration 3.93%). ).
(Iii) Preparation of composition After mixing the cherry extract and sake lees extract at a mixing ratio of 1: 9, the insoluble matter was filtered to obtain a brown transparent extract mixed solution (solid content concentration 0.31). %).

Preparation of Comparative Sample (1) A cherry extract solution was prepared by the same method as in Production Example 1 (i), and this was used as the comparative sample (1).

Preparation of Comparative Sample (2) A sake lees extract solution was prepared by the same method as in Production Example 2 (ii), and this was used as the comparative sample (2).

Prescription example 1. Toner [A component] Part Olive oil 1.0
Polyoxyethylene (5.5) cetyl alcohol 5.0
Butylparaben 0.1
[B component]
Composition of Production Example 1 (1) 5.0
Ethanol 5.0
Glycerin 5.0
1,3-butylene glycol 5.0
Potassium hydroxide Appropriate amount Purified water 100 parts in total [C component]
Appropriate amount of fragrance

Prescription example 2. Toner In the B component of Formulation Example 1, 5.0 parts of the composition (2) of Production Example 2 is used instead of the composition (1) of Production Example 1, and the lotion is prepared in the same manner as in Formulation Example 1. Obtained.

Prescription example 3. Toner In the B component of Formulation Example 1, 5.0 parts of the composition (3) of Production Example 3 is used instead of the composition (1) of Production Example 1, and the lotion is prepared in the same manner as in Formulation Example 1. Obtained.

Prescription example 4. Toner In the B component of Formulation Example 1, 5.0 parts of the composition (4) of Production Example 4 is used instead of the composition (1) of Production Example 1, and the lotion is prepared in the same manner as in Formulation Example 1. Obtained.

Prescription example 5. Toner In the B component of Formulation Example 1, 5.0 parts of the composition (5) of Production Example 5 is used instead of the composition (1) of Production Example 1, and the lotion is prepared in the same manner as in Formulation Example 1. Obtained.

Prescription example 6. Toner In the B component of Formulation Example 1, 5.0 parts of the composition (6) of Production Example 6 is used instead of the composition (1) of Production Example 1, and the lotion is prepared in the same manner as in Formulation Example 1. Obtained.

Prescription example 7. Toner In the B component of Formulation Example 1, 5.0 parts of the composition (7) of Production Example 7 is used instead of the composition (1) of Production Example 1, and the lotion is prepared in the same manner as in Formulation Example 1. Obtained.

Prescription example 8. Toner In the B component of Formulation Example 1, 5.0 parts of the composition (8) of Production Example 8 is used instead of the composition (1) of Production Example 1, and the lotion is prepared in the same manner as in Formulation Example 1. Obtained.

Prescription example 9. Toner In the B component of Formulation Example 1, 5.0 parts of the composition (9) of Production Example 9 is used instead of the composition (1) of Production Example 1, and the lotion is prepared in the same manner as in Formulation Example 1. Obtained.

Prescription example 10. Toner In the B component of Formulation Example 1, 5.0 parts of the composition (10) of Production Example 10 is used instead of the composition (1) of Production Example 1, and the lotion is prepared in the same manner as in Formulation Example 1. Obtained.

Prescription example 11. Toner In the B component of Formulation Example 1, 5.0 parts of the composition (10) of Production Example 11 is used instead of the composition (1) of Production Example 1, and the lotion is prepared in the same manner as in Formulation Example 1. Obtained.

Prescription example 12. Emulsion [Component A] Part Liquid paraffin 6.0
Hexalan 4.0
Jojoba oil 1.0
Polyoxyethylene (20) sorbitan monostearate 2.0
Soy lecithin 1.5
[B component]
Composition of Production Example 2 (2) 3.0
L-ascorbic acid-2-glucoside 2.0
Potassium hydroxide 0.5
Glycerin 3.0
1,3-butylene glycol 2.0
Carboxymethyl cellulose 0.3
Sodium hyaluronate 0.01
Amount of purified water totaling 100 parts [C component]
Appropriate amount of fragrance

Prescription example 13. Emulsion Emulsion in the same manner as in Formulation 12, except that 2.0 parts of tranexamic acid is used instead of 2.0 parts of L-ascorbic acid-2-glucoside and 0.5 part of potassium hydroxide in the B component of Formulation Example 12. Got

Prescription example 14. Emulsion Emulsion was prepared in the same manner as in Formulation 12 except that 3.0 parts of arbutin was used instead of 2.0 parts of L-ascorbic acid-2-glucoside and 0.5 part of potassium hydroxide in the B component of Formulation Example 12. Obtained.

Prescription example 15. Emulsion Same as in Formulation 12 except that 5.0 parts of nicotinamide is used instead of 2.0 parts of L-ascorbic acid-2-glucoside and 0.5 parts of potassium hydroxide in the B component of Formulation Example 12. Emulsion was obtained.

Prescription example 16. Essence [Ingredients] Part Ethanol 2.0
Glycerin 5.0
1,3-butylene glycol 5.0
Methylparaben 0.1
Hyaluronic acid 0.1
Composition of Production Example 5 (5) 5.0
Citric acid 0.3
Sodium citrate 0.6
Amount of purified water totaling 100 parts

Prescription example 17. Lotion [Ingredients] Part Composition of Production Example 6 (6) 10.0
Ethanol 10.0
Glycerin 3.0
1,3-butylene glycol 2.0
Methylparaben 0.2
Citric acid 0.1
Sodium citrate 0.3
Carboxyvinyl polymer 0.1
Xanthan gum 0.1
Perfume Appropriate amount Potassium hydroxide Appropriate amount Purified water 100 parts in total The above components were mixed to obtain a lotion.

Example 18. Liquid foundation [A component] Stearic acid 2.4
Propylene glycol monostearate 2.0
Setostearyl alcohol 0.2
Liquid lanolin 2.0
Liquid paraffin 3.0
Isopropyl myristate 8.5
Propylparaben 0.05
[B component]
Composition of Production Example 7 (7) 5.0
Sodium Carboxymethyl Cellulose 0.2
Bentonite 0.5
Propylene glycol 4.0
Triethanolamine 1.1
Methylparaben 0.1
Amount of purified water totaling 100 parts [C component]
Titanium oxide 8.0
Talc 4.0
Appropriate amount of coloring pigment

Prescription example 19. Hair shampoo [Ingredient A] Part N-coconut oil fatty acid methyl taurine sodium 10.0
Polyoxyethylene (3) Alkyl Ether Sodium Sulfate 20.0
Betaine Lauryl Dimethylamino Acetate 10.0
Coconut oil fatty acid diethanolamide 4.0
Methylparaben 0.1
[B component]
Citric acid 0.1
Composition of Production Example 10 (10) 2.0
1,3-butylene glycol 2.0
Amount of purified water totaling 100 parts

Example 20. Hair conditioner [Component A] Part Polyoxyethylene (10) Hardened castor oil 1.0
Distearyl dimethylammonium chloride 1.5
Stearyltrimethylammonium chloride 2.0
Glyceryl 2-ethylhexanoate 1.0
Cetanol 3.2
Stearyl alcohol 1.0
Methylparaben 0.1
[B component]
Composition of Production Example 1 (1) 2.0
1,3-butylene glycol 5.0
Amount of purified water totaling 100 parts

Prescription example 21. Hair tonic for hair growth [Ingredients] Part Dipotassium glycyrrhizinate 0.1
Mononitroguaiacol Sodium 0.02
Pyridoxine hydrochloride 0.03
Adenosine 1.0
Composition of Production Example 2 (2) 1.0
Trimethylglycine 0.5
Lactic acid 0.2
1,3-butylene glycol 10.0
Phenoxyethanol 0.2
Polyoxyethylene hydrogenated castor oil 0.4
Menispermaceae root extract 0.3
Panax ginseng extract 0.3
Appropriate amount of L-arginine Amount of purified water totaling 100 parts

The effectiveness of the composition of the present invention was evaluated by the following Test Examples 1 to 7, but the present invention is not limited to this.

Test example 1. ^{Evaluation of epidermal cell activating effect Human epidermal cell NHEK was seeded in 5 × 10 3} cells / hole in a 96-well microplate containing HuMedia KG2 medium (manufactured by Kurabo), and 1 under the conditions of 37 ° C and 5.0% CO _2. After pre-culturing for 1 day, a medium (Humedia KG2) containing each of the compositions (1) to (10) of Production Examples 1 to 10 as a sample solution was added to the pre-culture solution, and the cells were cultured under the same conditions for another 2 days. Here, the concentration of the sample solution was adjusted so that the final concentration as a solution with respect to the total amount of the medium to be added was 1.0% and 2.0%. After pre-culture, the medium is removed, 0.03% MTT is added and the temperature is kept at 37 ° C. for 1 hour, and then the produced formazan is extracted with isopropanol and used with a microplate reader (Model 680, manufactured by Biorad). The MTT value was measured at a wavelength of 570-630 nm. In addition, the same operation as above was performed in the case of no sample addition (Control). Then, the relative value of the MTT value at the time of adding each sample was obtained when the MTT value at the time of no sample addition was 100, and used as the epidermal cell MTT activity rate (%). A similar test was also performed when 100 mM glucose was added as a positive control instead of the sample solution to confirm that the test system was functioning normally.

The results of Test Example 1 are shown in Table 1.
[Table 1]

As shown in Table 1, it was confirmed that the compositions (1) to (10) according to the present invention have a remarkably excellent epidermal cell activating effect in a concentration-dependent manner. In addition, the same effect was obtained with glucose, confirming that this test system is functioning normally.

Test example 2. Evaluation of DPPH radical scavenging effect
2.4 parts of DPPH (1,1-diphenyl-2-picrylhydrazil) was dissolved in 20 parts of ethanol, and 20 parts of purified water was added thereto to prepare a DPPH solution. To 24 parts of this DPPH solution, 19.2 parts of 18v / v% ethanol solution and 4.8 parts of 2M sodium acetate-sodium acetate buffer (pH 5.5) were added to prepare a DPPH-added solution. In addition, in order to subtract the effect of the color tone of the extract itself on the test, a 50v / v% ethanol solution was used instead of the DPPH solution, and a mixture of 18v / v% ethanol solution and 2M sodium acetate-sodium acetate buffer was used. It was used as a control solution. Next, the compositions (1) to (10) of Production Examples 1 to 10 and the comparative samples (1) to (2) were diluted with purified water to prepare 12 kinds of sample solutions. Each composition was diluted with purified water so that the final concentration as a solution with respect to the total amount of the sample solution was 1.0% and 2.0%, respectively. This sample solution is mixed with the DPPH-added solution or the control solution at a ratio of 1: 3, and after allowing to stand at room temperature for 10 minutes, the absorbance at 550 nm when each sample solution is mixed with the DPPH-added solution and each sample solution are measured. The difference from the absorbance at 550 nm when mixed with the control solution was measured to confirm the residual amount of DPPH radicals. In addition, the same operation as above was performed in the case of no sample addition (Control), and the relative value of the DPPH radical residual rate at the time of adding each sample was obtained when the DPPH radical residual rate at the time of no sample addition was 100. .. A similar test was also performed when water-soluble vitamin E [Trolox] (final concentration 25 μM) was added as a positive control instead of the sample solution to confirm that the test system was functioning normally. ..

The results of Test Example 2 are shown in Table 2.
[Table 2]

As shown in Table 2, the compositions (1) to (10) according to the present invention have a concentration-dependently excellent DPPH radical scavenging action as compared with the comparative samples (1) and (2). Was shown. In addition, the same effect was obtained with water-soluble vitamin E [Trolox], confirming that this test system is functioning normally.

Test example 3. Evaluation of SOD-like action
1M Tris-hydrochloric acid buffer 0.15 mL, 1 mM ethylenediamine tetraacetic acid / disodium salt solution 0.30 mL, 1 mM xanthin solution 0.30 mL, 0.75 mM nitrobul-tetrazolium solution 0.20 mL, compositions of Production Examples 1 to 10 (1) to (10) ) And 0.10 mL of each of the comparative samples (1) and (2) and 1.90 mL of purified water were mixed to prepare 12 kinds of test solutions. In addition, a mixture (Control] having the same composition as the above test solution was prepared except that 2.00 mL of purified water was used instead of the test solution. Further, a mixed solution (positive control solution) having the same composition as the above test solution was prepared except that 0.10 mL of 0.875 Unit / mL superoxide dismutase (SOD) solution was used instead of the sample solution (0.10 mL). After incubating each of the above test solutions at 37 ° C, 0.05 mL of 1 Unit / mL xanthine oxidase solution was added thereto, and after a certain period of time (5 minutes), the absorbance of each test solution at 570 nm (superoxide in the test solution). An index of the amount of anion) was measured. The measurement results showed the absorbances of each test solution and positive control solution as relative values when the absorbance of the sample-free (Control) mixture was 100.

The results of Test Example 3 are shown in Table 3.
[Table 3]

As shown in Table 3, the compositions (1) to (10) according to the present invention have significantly superior SOD-like activity in a concentration-dependent manner as compared with the comparative samples (1) and (2). It has been shown. In addition, the same effect was obtained in SOD, which is a positive control, confirming that this test system is functioning normally.

Test example 4. ^{Evaluation of XVII type collagen gene expression Normal human epidermal cells were prepared in 6 × 10 5} cells / mL in HuMedia KG2 medium (manufactured by Kurabo) containing a growth additive, and 1 mL was seeded in a φ6 cm petri dish to generate 5% CO. _2. Incubated at 37 ° C under saturated steam. After culturing for 24 hours, the same medium containing the compositions (1) to (3) and (10) of Production Examples 1 to 3 and 10 as a sample solution (the final concentration as a solution is 1.0% with respect to the total amount of the medium). (The one containing the sample solution) was added and cultured. In addition, the same operation as above was performed in the case of no sample addition (Control). After culturing for 24 hours, cells in each test group were collected with 1 mL of Trizol reagent (manufactured by Invitrogen). Add 200 μL of chloroform (manufactured by Wako Pure Chemical Industries, Ltd.) to the collected cells, stir and mix, and centrifuge with a centrifuge (manufactured by TOMY / MX-160) at 15,000 rpm for 15 minutes at 4 ° C. After that, only the aqueous layer was separated by 400 μL. 500 μL of isopropanol (manufactured by Wako Pure Chemical Industries, Ltd.) was added to the recovered aqueous layer, stirred and mixed, and centrifuged at 15,000 rpm and 4 ° C. for 15 minutes to obtain a total RNA precipitate. 1 mL of 75% ethanol was added to total RNA, stirred and washed, and centrifuged at 15,000 rpm for 15 minutes at 4 ° C. to recover the precipitate. The collected total RNA was reverse-transcribed using a predetermined kit (PrimeScript RT reagent Kit with gDNA Eraser (Perfect Real Time) [manufactured by Takara Bio Inc.]) to synthesize cDNA. Using the synthesized cDNA as a sample, using Thermal Cycler Dice (registered trademark) Real Time System Single (manufactured by Takara Bio) and SYBR (registered trademark) Premix Ex TaqTM II (Perfect Real Time) [manufactured by Takara Bio]. The expression of the XVII type collagen gene and the expression of the internal standard β-actin gene were detected. Here, β-actin is one of the housekeeping genes (genes that are commonly expressed in many tissues and cells in a certain amount, are always expressed, and are indispensable for cell maintenance and proliferation). Since the expression level is always constant, it is used as an internal standard in PCR experiments. The test results compared the expression levels of the XVII collagen gene in each test group when the expression level of the β-actin gene was constant. In this test system, the relative value of the expression level of the gene in the test group was determined when the expression level of each gene in the control group was 100.

The results of Test Example 4 are shown in Table 4.
[Table 4]

As shown in Table 4, the composition according to the present invention was shown to significantly induce gene expression of type XVII collagen.

Test example 5. XVII type collagen synthesis promoting effect Normal human skin-derived epidermal cells (NHEK) were seeded in 96-well microplates containing HuMedia KG2 medium (manufactured by Kurabo) at 8 × 10 ³ cells / hole at 37 ° C, 5.0% CO ₂ . After pre-culturing under the conditions for 1 day, the same medium containing the sample solution (the one containing the sample solution so that the final concentration as a solution is 1.0% or 2.0% with respect to the total amount of the medium) is added and added. The cells were cultured for another 2 days under the conditions. After that, immunological detection was performed using a type XVII collagen antibody. That is, after washing with PBS (-), cells were treated with 15% neutral buffered formalin solution for 30 minutes to fix them, permeated with 0.5% Triton X-100 solution for 1 hour, and 5-fold diluted blocking one P (Nacalai Tesque). After blocking by treatment with a solution for 2 hours, type XVII collagen antibody was added, and the mixture was allowed to stand at 4 ° C. for 24 hours. Then, it was washed with PBS (-), a fluorescently labeled secondary antibody was added, and the mixture was allowed to stand in a dark place for a certain period of time. After the PBS (-), it was washed and the fluorescence intensity was measured. First, the fluorescent label (Alexa Fluor488) of the secondary antibody was measured at Ex = 485 nm and Em = 520 nm [fluorescent microplate reader (Fluoroscan Ascent, manufactured by Thermo Fisher Scientific)], and then DNA staining with Hoechst33342 was performed. Measurements were performed at Ex = 355 nm and Em = 460 nm. The degree of XVII collagen production was determined by dividing the fluorescence intensity of Alexa Fluor 488 in each test group by the fluorescence intensity of Hoechst 33342. In addition, in the case of no sample addition (Control), the same operation as above is performed, and the relative value of the XVII type collagen production degree at the time of adding each sample to the XVII type collagen production degree obtained here is obtained, and the XVII type collagen is obtained. The amount produced (%) was used.

The results of Test Example 5 are shown in Table 5.
[Table 5]

As shown in Table 5, it was confirmed that the composition according to the present invention has an excellent concentration-dependent type XVII collagen synthesis promoting action. Type XVII collagen plays a role of binding the basement membrane and the epidermis existing at the junction between the epidermis and the dermis, and the basement membrane plays a role of maintaining the expression of differentiation traits of the basal cells. , It is suggested that by inducing the expression of type XVII collagen, it has the effect of maintaining the basement membrane in a normal state and the skin metabolism (turnover) in a normal state. It is also suggested that by inducing the expression of type XVII collagen in the scalp, it has an effect of preventing hair loss and gray hair.

Test example 6. Suppressive effect on melanin synthesis Cultured B16 mouse melanoma cells B16-F10 were ^{seeded in a 24-well microplate at 2.4 × 10 4} cells / well, and were seeded in RPMI1640 medium containing 10% FBS for 1 day under the conditions of _{37 ° C and 5.0% CO 2.} After pre-culture, the sample solution was used as a solution, diluted to a final concentration of 1.0% and 2.0%, and a theophylline-containing medium adjusted to a final concentration of 1 mM was added, and under the same conditions, 2 Incubated for days. The culture was then removed, 200 μL of 1N NaOH / 10% dimethylsulfoxide solution was added per hole, sealed and incubated at 50 ° C. for 2 hours to lyse the cells. 100 μL of this solution was transferred to another 96-well microplate, and the melanin level was measured at a wavelength of 490 nm using a microplate reader (Model 680, manufactured by Bio-Rad). On the other hand, 5 μL of the solution in which the same cells were lysed was transferred to another 96-well microplate, and 200 μL of Dye Reagent Concentrate (Biorad) solution diluted 5-fold with purified water was added, and a microplate reader (Model 680, Biorad) was added. The absorbance at a wavelength of 570 nm was measured using (manufactured by the same company). Another known amount of bovine serum albumin (manufactured by Sigma) was serially diluted, and the amount of protein equivalent to albumin was measured from the calibration curve obtained by the same operation. The obtained absorbance was divided by the amount of protein to determine the amount of melanin per protein. Further, in the case of no sample addition (Control), the same operation as above is performed, and the relative value of the melanin amount per protein at the time of adding each sample to the melanin amount per protein obtained here is obtained, and the melanin synthesis rate is obtained. It was set to (%). For comparison, the same test was performed when 2 mM kojic acid was added instead of the sample solution (positive control).

The results of Test Example 6 are shown in Table 6.
[Table 6]

As shown in Table 6, the composition according to the present invention has a remarkably excellent melanin synthesis inhibitory effect, suggesting a preventive and ameliorating effect on spots and freckles.

Test example 7. Suppression of Perilipin Synthesis Hereinafter, the effect of suppressing fat accumulation of the composition according to the present invention will be evaluated based on the effect of suppressing the synthesis of the protein [perilipin 1] present around the lipid droplets in adipocytes. Since it is known that when the amount of perilipin 1 decreases, the number of fat droplets decreases and the size of lipid droplets decreases, the effect of suppressing the synthesis of perilipin 1 is evaluated.
^{Mouse fibroblasts (3T3-L1) were seeded at 1.5 x 10 4} cells / hole in a 96-well microplate containing 10% FBS-containing Dalveco modified Eagle's minimum essential medium (DMEM: Nissui Pharmaceutical Co., Ltd.), 37 After preculture for 3 days under the conditions of ℃ and 5.0% CO ₂ , differentiation-inducing medium (10% FBS, 0.5 mM 3-isobutyl-1-methylxanthin (IBMX), 0.25 μM dexamethasone (DEX) and 1.1 μg / mL insulin) After adding DMEM) mixed with, the cells were cultured for another 2 days. Then, the same medium containing the sample solution (the one containing the sample solution so that the final concentration as a solution was 1.0% and 2.0% with respect to the total amount of the medium) was additionally added, and the cells were cultured under the same conditions for another 5 days. Then, immunodetection was performed using a perilipin1 antibody. That is, after washing with PBS (-), cells were treated with 10% neutral buffered formalin solution for 30 minutes to fix, permeated with 0.5% Triton X-100 solution for 1 hour, and 5-fold diluted Blocking One P (Nacalai Tesque). ) After blocking by treatment with the solution for 2 hours, perilipin1 antibody was added, and the mixture was allowed to stand at room temperature for 1 hour. Then, it was washed with PBS (-), a fluorescently labeled secondary antibody was added, and the mixture was allowed to stand in a dark place for a certain period of time. After that, it was washed with PBS (-) and the fluorescence intensity was measured. First, the fluorescent label (Alexa Fluor488) of the secondary antibody was measured at Ex = 485 nm and Em = 520 nm [fluorescent microplate reader (Fluoroscan Ascent, manufactured by Thermo Fisher Scientific)], and then DNA staining with Hoechst33342 was performed. Measurements were performed at Ex = 355 nm and Em = 460 nm. The degree of perilipin1 formation was determined by dividing the fluorescence intensity of Alexa Fluor 488 in each test group by the fluorescence intensity of Hoechst 33342. In addition, in the case of no sample addition (Control), the same operation as above was performed, and the relative value of the perilipin1 formation degree at each sample addition was obtained with respect to the perilipin1 formation degree obtained here, and the perilipin1 synthesis rate (%) was obtained. did.

The results of Test Example 7 are shown in Table 7.
[Table 7]

As shown in Table 7, it was confirmed that the composition according to the present invention has a remarkably excellent effect of suppressing perilipin synthesis in a concentration-dependent manner.

Claims (3)

An external preparation for skin containing an extract of a cherry flower belonging to the genus Prunus of the Rosaceae family and an extract of yeast. An external preparation for the skin containing an extract of cherry blossoms belonging to the genus Prunus of the Rosaceae family and an extract of sake lees. An external preparation for skin containing an extract of a cherry flower belonging to the genus Prunus of the Rosaceae family and a fermented product of rice yeast and aspergillus.